In both plant and animal systems, a central question of stem cell biology is how stem cell populations are established and then maintained long term. Our studies aim to address this central question. One of the most important events during plant embryogenesis involves formation of the root meristem. The root meristem is derived from a single cell, referred to as the hypophyseal cell, early in embryogenesis and consists of the quiescent center (QC) surrounded by distal and proximal stem cell populations. Our studies point to a mechanism whereby an interplay between hormone signaling and transcription factors creates and maintains the distal stem cell population. Understanding the logic between this interplay of dynamic signaling and stable establishment of stem cell fate is likely to have broad applications in both the agricultural and health sciences. We previously demonstrated that the NO TRANSMITTING TRACT (NTT) gene encodes a transcription factor that is required for normal carpel and fruit development. The current proposal focuses on our recent discovery that mutations in NTT, when combined with mutations in the two most closely related genes, WIP4 and WIP5, abolish root meristem formation. Our studies suggest that NTT, WIP4 and WIP5 (hereafter, the NWW genes) act redundantly and are both necessary and sufficient to specify distal stem cell fate during embryogenesis. Importantly, all three genes are expressed early in embryo development in the hypophyseal cell, and this expression pattern is dependent on the auxin-signaling pathway. Our studies directly refute widely accepted models and provide a simple hypothesis to explain how the plant hormone auxin patterns stem cell identity. Our studies suggest that one of the critical steps in stem cell patterning during embryogenesis involves the precise temporal and spatial pattern of NWW gene expression. We propose to test this hypothesis in order to provide a mechanistic understanding of stem cell specification. Because of their critical role in meristem patterning, we propose to use the NWW genes as a platform to identify the key factors that specify and maintain stem cell fate in embryonic and root meristems. These factors include the upstream regulators of NWW gene expression as well as the factors that interact with and/or function downstream of NWW. We propose a series of molecular and genetic approaches to identify and characterize these factors in order to provide a framework for future efforts aimed at understanding stem cell specification and maintenance.